Genetic lineage tracing of resident stem cells by DeaLT
Genetic lineage tracing of resident stem cells by DeaLT
复制标题
通过 DeaLT 对驻留干细胞进行遗传谱系追踪
DOI：
10.1038/s41596-018-0034-5
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发表时间：
2018-10-01
期刊：
影响因子：
14.8
通讯作者：
Zhou, Bin
中科院分区：
文献类型：
--
作者：
He, Lingjuan;Li, Yan;Zhou, Bin
揭示驻留干细胞在组织再生过程中的命运是临床和基础研究的重要目标。基于Cre-loxP重组的遗传谱系追踪为推断体内细胞命运和细胞转化提供了有效的策略。然而，在疾病状态下和组织再生期间确定驻留干细胞或其衍生物的确切命运在许多研究领域中仍然存在争议，部分原因是与基于Cre的谱系追踪相关的技术限制，例如脱靶标记。最近，我们生成了一个新的谱系追踪平台，我们将其命名为DeaLT（双重组酶激活谱系追踪），该平台使用Dre-rox重组系统来提高Cre介导的谱系追踪的精度。在这里，我们描述了一个详细的协议，使用DeaLT跟踪c-Kit（+）心脏干细胞及其衍生物的命运，在没有任何干扰的非靶细胞，如心肌细胞，在器官稳态和组织损伤后。这种谱系追踪方案也可用于描绘其他器官系统的常驻干细胞的命运，从小鼠杂交到最终组织分析，需要大约10个月的时间才能完成。
Unraveling the fates of resident stem cells during tissue regeneration is an important objective in clinical and basic research. Genetic lineage tracing based on Cre-loxP recombination provides an effective strategy for inferring cell fate and cell conversion in vivo. However, the determination of the exact fates of resident stem cells or their derivatives in disease states and during tissue regeneration remains controversial in many fields of study, partly because of technical limitations associated with Cre-based lineage tracing, such as, for example, off-target labeling. Recently, we generated a new lineage-tracing platform we named DeaLT (dual-recombinase-activated lineage tracing) that uses the Dre-rox recombination system to enhance the precision of Cre-mediated lineage tracing. Here, we describe as an example a detailed protocol using DeaLT to trace the fate of c-Kit(+) cardiac stem cells and their derivatives, in the absence of any interference from nontarget cells such as cardiomyocytes, during organ homeostasis and after tissue injury. This lineage-tracing protocol can also be used to delineate the fate of resident stem cells of other organ systems, and takes similar to 10 months to complete, from mouse crossing to final tissue analysis.